Estrogens play an important role in the phenotypic expression of their target tissues, where they affect cellular growth, differentiation, and malignancy. To date, little has been done to examine thenegative effects of this hormone on the expression of specific genes. In this proposal we will target the synthesis of serum albumin in Xenopus laevis as a potential model system for the negative regulation of gene expression by estrogen. The administration of estrogen to male or female Xenopus induces the synthesis of large quantities of the yolk protein precursor vitellogenin and appears to suppress the constitutive synthesis of serum albumin. We will examine this phenomenon in detail through the use of specific antibodies and recombinant DNA probes for both albumin and vitellogenin in order to determine if there indeed is differential regulation of these genes or whether albumin gene expression remains constitutive throughout the vitellogenic response. In the initial experiments we will study the effects of estrogen on the relative rates of albumin and virellogenin synthesis, and concomitantly assess changes in the amount of translatable mRNA for each protein by in vitro translation in a reticulocyte lysate. We have cloned the structural genes for serum albumin and have found that this protein is encoded by 2 mRNAs. The cDNA clones will be sequenced to assess their relationships to each other and to mammalian albumin and alphafetoprotein. We will use these clones and a series of virellogenin clones that we have obtained to study the effects of estrogen on the steady state levels of albumin and vitellogenin mRNA in vivo. We will then turn to an in vitro organ culture system in whih we will perform pulse-labeling experiments to assess the effects of estrogen on the rates of transcription and degradtion of albumin and vitellogenin mRNA. The possibility of differential regulation by estrogen of the members of the albumin gene family will be considered in both the in vivo experiments. In this project we will also use the cDNA clones for serum albumin to isolate the chromosomal genes from a cloned library of Xenopus DNA. The structural relationships of these genes will be analyzed by heteroduplex, R-loop and restriction mapping.